Transcriptomics

Dataset Information

3

Interaction with Human Bronchial Epithelial Cells in vitro Affects Gene Expression in Moraxella catarrhalis


ABSTRACT: In this study, DNA microarray technology was used to measure global gene expression in M. catarrhalis cells that had attached to human bronchial epithelial cells in culture. M. catarrhalis ATCC 43617 was allowed to attach (MOI~30) to a monolayer of 16HBE14 human bronchial epithelial cells for 2 hr in OmniTray cell culture plates. Additional portions of these bacteria were incubated in tissue culture medium in 75-cm2 cell culture flasks in the absence of 16HBE14 cells for 2 hr. After this incubation period, the medium in the flask with the 16HBE14 monolayer was decanted and replaced with tissue culture medium containing 25 mM sodium azide (to protect RNA). The 16HBE14 monolayer and attached bacteria were released from the plastic by scraping. After low-speed centrifugation to collect these bronchial cells and attached bacteria, the cell pellet was re-suspended in TE containing 1% saponin (which lyses the human cells). After a 10-min incubation at 37°C, the bacterial cells were collected by centrifugation. The bacteria growing in the flask without human cells were treated identically and then harvested by centrifugation. Total RNA was extracted from both sets of bacterial cells by using the Qiagen RNeasy Midi kit. DNA microarray analysis, slide scanning, and statistical analysis of the data were performed. RNA was isolated four times and used in five DNA microarray hybridizations; “dye swap” experiments were performed twice. Two additional independent experiments were performed to isolate two sets of RNA samples for qRT-PCR analysis to validate DNA microarray data.

ORGANISM(S): Moraxella catarrhalis  

SUBMITTER: Eric J Hansen   Wei Wang  Eric Hansen 

PROVIDER: E-GEOD-12990 | ArrayExpress | 2014-03-24

SECONDARY ACCESSION(S): GSE12990PRJNA112599

REPOSITORIES: GEO, ArrayExpress

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Publications

Identification of an outer membrane lipoprotein involved in nasopharyngeal colonization by Moraxella catarrhalis in an animal model.

Wang Wei W   Joslin Stephanie N SN   Pybus Christine C   Evans Amanda S AS   Lichaa Flora F   Brautigam Chad A CA   Hansen Eric J EJ  

Infection and immunity 20140318 6


Colonization of the human nasopharynx by Moraxella catarrhalis is presumed to involve attachment of this bacterium to the mucosa. DNA microarray analysis was used to determine whether attachment of M. catarrhalis to human bronchial epithelial (HBE) cells in vitro affected gene expression in this bacterium. Attachment affected expression of at least 454 different genes, with 163 being upregulated and 291 being downregulated. Among the upregulated genes was one (ORF113) previously annotated as enc  ...[more]

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